Given the growing need for replacement tissues, organs and grafts, tissue engineering has emerged as a viable regenerative medicine technology. Recent developments in the field suggest that it is possible to fabricate functional organ modules under laboratory conditions and these results may soon translate to clinically relevant treatment options. The present Phase I application describes an approach for clinically appropriate cell sourcing options to be combined with cutting-edge bioprinting technology for the fabrication of vascular tissues. While it is desirable to fabricate tissue engineered blood vessels utilizing cells derived directly from native blood vessels, it is unlikely that sufficient numbers of cells will be available for procurement from vasculopathic patients. We, therefore, propose to isolate endothelial cells (EC) and multipotential stromal/stem cells (MSC) from the stromal vascular fraction (SVF) of human adipose tissue, which is both abundant and easily accessible. Isolated MSC will be differentiated in vitro into smooth muscle cells (SMC) that can then be combined with EC isolated from the original adipose tissue specimen for the purpose of bioprinting. Classical methods of tissue engineering rely on cell seeding or sodding of scaffold materials, which are either degraded or permanently incorporated into the tissue construct. The presence of scaffold material represents potential difficulties with respect to particle remnant during degradation and sites for potential infection. To overcome the limitations of the classical approach we have implemented a patent- pending "print-based" tissue engineering technology. Bioprinted vessel constructs will be fabricated using Organovo, Inc.'s patent-pending technology involving the production of cylindrical cellular intermediates which can be bioprinted in the context of supporting hydrogel materials to form 3-D blood vessel constructs of varying dimensions. This method of tissue engineered blood vessel fabrication results in a scaffold-free bioengineered blood vessel, which can be further matured under pulsatile flow conditions in a perfusion bioreactor to yield blood vessel constructs with appropriate cellular and matrix compositions resulting in improved biomechanical characteristics. The principal objective of this project is to develop a clinically applicable strategy for sourcing the cellular material needed for fabrication of tissue engineered blood vessels and to successfully develop such tissues using Organovo's methods of bioprinting. Completion of this project represents a critical component in the development of clinically available bioengineered tissues. PUBLIC HEALTH RELEVANCE: The objective of this proposal is to construct biologically relevant small diameter blood vessels using a clinically relevant cell source. Cells from the stromal vascular fraction of adipose tissue will be incorporated into vascular conduits made with a demonstrated bioprinting method, and their biomechanical properties and structure will be assessed. The proposed studies could lead to implantable blood vessels that mimic native arteries and are composed of the patient's own cells (an autologous model).